The TPM3‐NTRK1 rearrangement is a recurring event in colorectal carcinoma and is associated with tumor sensitivity to TRKA kinase inhibition

Abstract

Growth Factor Receptor. NTRK1 was originally isolated from a colorectal carcinoma (CRC) sample as component of a somatic rearrangement (TPM3-NTRK1) resulting in expression of the oncogenic chimeric protein TPM3-TRKA, but there has been no subsequent report regarding the relevance of this oncogene in CRC. The KM12 human CRC cell line expresses the chimeric TPM3-TRKA protein and is hypersensitive to TRKA kinase inhibition.We report the detailed characterization of the TPM3-NTRK1 genomic rearrangement in KM12 cells and through a cellular screening approach, the identification of NMS-P626, a novel highly potent and selective TRKA inhibitor. NMS-P626 suppressed TPM3-TRKA phosphorylation and downstream signaling in KM12 cells and showed remarkable antitumor activity in mice bearing KM12 tumors.Finally, using quantitative reverse transcriptase PCR and immunohistochemistry (IHC) we identified the TPM3-NTRK1 rearrangement in a CRC clinical sample, therefore suggesting that this chromosomal translocation is indeed a low frequency recurring event in CRC and that such patients might benefit from therapy with TRKA kinase inhibitors.

The molecular landscape of colorectal cancer cell lines unveils clinically actionable kinase targets

“…In some instances (FGFR2, ALK or NTRK1), overexpression of the TK gene was associated with amplification or translocation of the corresponding locus. The same genetic alterations have been previously reported in CRC cells and patients [26][27][28] , thus validating our approach. Our data further suggest that overexpression of TK sustains primary resistance to EGFR blockade, and could be used to identify patients unlikely to respond to cetuximab or panitumumab.…”

“…We decided to focus on TKs for which targeted agents are in clinical trial or are already approved for treatment, namely ALK, FGFR2, KIT, NTRK1, NTRK2, RET and PDGFRA. This analysis revealed that the overexpression of NTRK1 and FGFR2 is associated to molecular alterations, such as gene translocation (NTRK1) or gene amplification (FGFR2; Supplementary Figs 5 and 6), previously described in cellular models and in cancer patients [26][27][28] , thus validating the experimental approach. The other TK outlier genes were not previously reported in CRC cells.…”

Entrectinib, a Pan–TRK, ROS1, and ALK Inhibitor with Activity in Multiple Molecularly Defined Cancer Indications

“…However, chromosomal rearrangements involving the NTRK1 gene, resulting in the expression of different TRKA fusion proteins were recently reported by Vaishnavi and colleagues (11) in NSCLC, along with robust preclinical demonstration of the oncogenic potential of the predicted chimeric proteins. In addition, we very recently reported the presence of the recurring rearrangement TPM3-NTRK1 in colorectal carcinoma patients, together with preclinical validation data supporting the role of TPM3-TRKA fusion protein as a driver for proliferation and survival of TRKA-positive tumors (12).…”

“…The antiproliferative activity of test compounds and the effect of entrectinib on cell cycle were evaluated as previously described (12). Western blot analysis Cellular mechanism of action of entrectinib was investigated as described (12). The following antibodies were used for Western blot analysis: Phospho-TRKA-Tyr490 (#625725) and TRKA (#PC31) from Calbiochem, ALK (#35-4300) from Invitrogen, Phospho-ALK-Tyr1604 (#3341), Phospho-ROS1-Tyr2274 (#3078), ROS1 (#3266), Phospho-PLCg1-Tyr783 (#2821), PLCg1 (#2822), Phospho-STAT3-Tyr705 (#9131), Phospho-AKT-Ser473 (#9271), AKT (#9272), Phospho-MAPK-Thr202/Tyr204 (#9101), MAPK (#9102), caspase-3 (#9662), cleaved caspase-3 (Asp175; #9661), and PARP (#9542) from Cell Signaling Technology, STAT3 (#610189), p21 (#556431) and p27 (#610242) from BD Biosciences, actin (#A4700) from SIGMA, GAPDH (#sc-25778) from Santa Cruz Biotechnology.…”

“…The recent description of recurring rearrangements of the NTRK1 gene, which encodes this kinase, in subsets of major indications such as NSCLC, colorectal carcinoma together with the identification of NTRK2 and NTRK3 rearrangements in glial tumors and inflammatory breast cancer, respectively, and perhaps additional tumor types (11,12,14,16,17), has highlighted the relevance of these kinases as pharmacologic targets and the clinical need for effective inhibitors.…”